The utilization of phosphate ions from hydroxyapatite (tooth enamel) by oral bacteria may be significant in the carious process. Single and dual in vivo labeling of rat enamel (Ca45 and P32) will provide a method for studying the fate of these ions in an in vitro culture system designed so that cariogenic bacteria can utilize enamel phosphate as substrate. Individual components of the culture system will be analyzed to determine the disposition of the calcium and phosphate ions during bacterial metabolism. An isolate from deep dentinal caries (L. casei) that forms large intracellular granules will be used as a prototype for these experiments. In proposed studies, the optimum growth conditions for the formation of these granules will be determined using a relatively defined medium and varying nutritional and physical parameters. After these optimums are established, Ca 45 and/or P32 labeled hydroxyapatite will be substituted for phosphate in the medium. When growth is completed, the supernatant fluid, the individual cells and isolated granules will be analyzed for labeled ions. In addition, the chemistry and structure of the granules will be determined, the transferring enzymes identified and the effect of inhibitors examined. The methods of procedure developed with the prototype organism will be used to study strains of S. mutans and S. salivarius described as cariogenic by other laboratories.